A.Cavallaro at botany.uq.oz.au (Tony Cavallaro) wrote:
>In out lab lately we have bben trouble growing up DH5alpha cell
>after they have been glycerol solution frozen at -80 degrees. They grow
>fine in LB-amp (50), but fail to give any DNA after a miniprep procedure.
>THis is unusual since we usually have no trouble with this.
>>Also another problem with minipreps, we are also having trouble growing
>up colonies from plates (both LB agar and Mcconkey). The colonies look
>good on the plate but do not grow up in liquid culture. We are using
>DH5alpha again for this.
>>Help for this problem would be appreciated.
>>Tony
We've had this problem with this strain in particular. Run a sample
of the miniprep uncut (i.e. never seen Mg++) . In our case, this
showed DNA --- add Mg++ and it dissapears. Nuclease contamination
seems to be the culprit. We used an additional gene clean step to
clean the DNA up a little more.
Yours,
John R. Thompson
Merck Research Labs